Multidrug resistance protein-mediated transport of chlorambucil and melphalan conjugated to glutathione.
The human multidrug resistance protein (MRP1) confers resistance of cells to a number of different cytostatic drugs and functions as an export pump for glutathione S-conjugates, glucuronides and other amphiphilic anions. The present study details for the first time MRP1-mediated ATP-dependent transport of various glutathione S-conjugates of the bifunctional alkylating agents chlorambucil and melphalan. In membrane vesicles prepared from cells expressing recombinant MRP1, the conjugates were transported at rates in the following order: monoglutathionyl chlorambucil > bisglutathionyl chlorambucil > monohydroxy monoglutathionyl chlorambucil and monoglutathionyl melphalan > monohydroxy monoglutathionyl melphalan. In addition, we show that membranes from chlorambucil-resistant GST-alpha-overexpressing CHO cells as well as from their parental cells express the hamster homologue of MRP1. With both CHO cell membrane preparations, we observed ATP-dependent transport of monoglutathionyl chlorambucil and of leukotriene C4, a glutathione S-conjugate and high-affinity substrate of MRP1. The transport rates measured in the resistant cells were only two- to three-fold higher than those measured in the control cells. These results together with cytotoxicity assays comparing MRP1-overexpressing cell pairs with the CHO cell pair indicate that, although MRP1-mediated transport is active, it may not be the rate-limiting step in chlorambucil resistance in these cell lines.
Barnouin
K
K
Deutsches Krebsforschungszentrum, Heidelberg, Germany.
Leier
I
I
Jedlitschky
G
G
Pourtier-Manzanedo
A
A
König
J
J
Lehmann
W D
WD
Keppler
D
D
D000595:Amino Acid Sequence; D000818:Animals; D018906:Antineoplastic Agents, Alkylating / Q000378:metabolism*; D001693:Biological Transport, Active; D019328:Buthionine Sulfoximine / Q000494:pharmacology; D016466:CHO Cells; D002462:Cell Membrane / Q000378:metabolism; D002699:Chlorambucil / Q000378:metabolism*; D006224:Cricetinae; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004351:Drug Resistance; D005978:Glutathione / Q000031:analogs & derivatives* / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D017997:Leukotriene C4 / Q000378:metabolism; D013058:Mass Spectrometry / Q000379:methods; D008558:Melphalan / Q000378:metabolism*; D008969:Molecular Sequence Data; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D000074081:MutS Homolog 3 Protein; D016415:Sequence Alignment; D014407:Tumor Cells, Cultured
D018906:Antineoplastic Agents, Alkylating; D019328:Buthionine Sulfoximine; D002699:Chlorambucil; D004268:DNA-Binding Proteins; D005978:Glutathione; D017997:Leukotriene C4; D008558:Melphalan; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D000074081:MutS Homolog 3 Protein; C078444:MSH3 protein, human; C421109:multidrug resistance-associated protein 1